Structure of the Polyadenylation Regulatory Element of the Human U1A Pre-mRNA 3‘-Untranslated Region and Interaction with the U1A Protein

Gubser, Charles C.; Varani, Gabriele

doi:10.1021/bi952319f

Cited by 75 publications

(87 citation statements)

References 48 publications

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“…Since C-SLC3 and C-SLC5 have different conformations based on nondenaturing gel electrophoresis, and since both are recognized by the CMV replicase, replicase-promoter recognition likely takes place by an induced-fit mechanism whereby the replicase and/or the RNA undergoes conformational changes after the initial interaction (20). Interaction of the BMV replicase with the CMV core elements in the CMV tRNA-like structure also likely takes place by an induced-fit mechanism.…”

Section: Discussionmentioning

confidence: 99%

Recognition of the Core RNA Promoter for Minus-Strand RNA Synthesis by the Replicases of Brome Mosaic Virus and Cucumber Mosaic Virus

Sivakumaran

Bao

Roossinck

et al. 2000

J Virol

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Replication of viral RNA genomes requires the specific interaction between the replicase and the RNA template. Members of the Bromovirus and Cucumovirus genera have a tRNA-like structure at the 3 end of their genomic RNAs that interacts with the replicase and is required for minus-strand synthesis. In Brome mosaic virus (BMV), a stem-loop structure named C (SLC) is present within the tRNA-like region and is required for replicase binding and initiation of RNA synthesis in vitro. We have prepared an enriched replicase fraction from tobacco plants infected with the Fny isolate of Cucumber mosaic virus (Fny-CMV) that will direct synthesis from exogenously added templates. Using this replicase, we demonstrate that the SLC-like structure in Fny-CMV plays a role similar to that of BMV SLC in interacting with the CMV replicase. While the majority of CMV isolates have SLC-like elements similar to that of Fny-CMV, a second group displays sequence or structural features that are distinct but nonetheless recognized by Fny-CMV replicase for RNA synthesis. Both motifs have a 5CA3 dinucleotide that is invariant in the CMV isolates examined, and mutational analysis indicates that these are critical for interaction with the replicase. In the context of the entire tRNA-like element, both CMV SLC-like motifs are recognized by the BMV replicase. However, neither motif can direct synthesis by the BMV replicase in the absence of other tRNA-like elements, indicating that other features of the CMV tRNA can induce promoter recognition by a heterologous replicase.An essential feature of viral infection is the replication of the viral genome. Many of the plant and animal viruses have plusstrand RNA genomes that, following entry into cells, can readily be translated to provide the proteins required for their replication. In addition, the genomic RNA of these viruses directs the synthesis of a complementary minus strand, which serves as the template for synthesis of multiple copies of genomic and possible subgenomic plus-strand RNA (8). Brome mosaic virus (BMV) and Cucumber mosaic virus (CMV) are the type members of the Bromovirus and Cucumovirus genera, respectively, in the family Bromoviridae, which belongs to the alpha-like superfamily of viruses (19,28). CMV and BMV have a similar genome organization, but CMV has a broader host range and a more severe economic impact (3,29,35). Both viruses have tripartite genomes, designated RNA1, RNA2, and RNA3. RNA1 encodes the putative protein with helicase and RNA-capping activities (5, 27), while RNA2 encodes the RNA-dependent RNA polymerase (RdRp). RNA3 is dicistronic and encodes the movement protein and the coat protein that is expressed via a subgenomic RNA. In Cucumoviruses but not Bromoviruses, an additional subgenomic RNA, 4a, that corresponds to the 3Ј-proximal portion of RNA2 has also been identified (13,35). RNA4a encodes the 2b protein and may be involved in systemic infection, pathogenicity, and suppressing posttranscriptional gene silencing (7,14,15,30).Replication of the BMV and CMV ...

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Section: Discussionmentioning

confidence: 99%

Recognition of the Core RNA Promoter for Minus-Strand RNA Synthesis by the Replicases of Brome Mosaic Virus and Cucumber Mosaic Virus

Sivakumaran

Bao

Roossinck

et al. 2000

J Virol

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“…The RNA spectra refer to a 30-nt construct of the polyadenylation regulatory element of human U1A pre-mRNA 3 0 untranslated region (3 0 UTR RNA). 44 The figures depicted are superpositions of two samples partially labeled with guanine and uracyl (purple) or with cytosine and adenine (green/ orange); partial labeling reduces spectral overlap for the RNA, allowing many more sites to be sampled. The DNA spectrum refers to a uniformly labeled sample of the binding site for HhaI Methyltransferase.…”

Section: Nmr Studies Of Fast Molecular Motions By Relaxation Rate Meamentioning

confidence: 99%

NMR studies of dynamics in RNA and DNA by ¹³C relaxation

Shajani

Varani

2006

Biopolymers

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T he many functions of RNA in biology very often require this molecule to change its conformation in response to biological signals in the form of small molecules, proteins, or other nucleic acids. 1,2 Although structurally more conservative and therefore dynamically less diverse, local motions in DNA may facilitate protein recognition and allow enzymes acting on DNA to access functional groups on the bases that would otherwise be buried in Watson-Crick base pairs. 3 These statements make a compelling case to study the sequence dependent dynamics in nucleic acids; yet, residue-specific studies of nucleic acid dynamics are relatively few. Fortunately, NMR studies of dynamics of nucleic acids and nucleic acids-protein complexes are gaining increased attention, this spectroscopy being the only way to access information on a residue-by-residue basis. The questions that are being asked are how motion contributes to the affinity and specificity of biological interactions and what trajectories lead to the remarkably large conformational changes that are sometimes observed.RNA and DNA molecules experience motions on a wide range of time scales, ranging from rapid localized motions to much slower collective motions of entire helical domains. Figure 1 summarizes the NMR spectroscopic techniques commonly used to obtain information in each motional regime. Dynamics occurring on time scales that are considered ''fast'' in NMR spectroscopic studies (ps-ns) largely reflects bond librations associated with small amplitude and localized motions.

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“…Previously, interactions such as Tat-TAR interaction of HIV-1 or the binding of U1A protein to its RNA substrate have been assigned to the induced fit category (Puglisi et al 1992(Puglisi et al , 1995Oubridge et al 1994;Avis et al 1996;Gubser and Varani 1996;Aboul-ela and Varani 1998;Varani et al 2000;Reyes et al 2001). In these cases the conformation of the unbound RNA is stable but significantly differs from that of the bound RNA.…”

Section: Protein-assisted Rna Foldingmentioning

confidence: 99%

“…In the induced fit, the RNA undergoes a transition between two different well-defined conformations, whereas conformational capture refers to the stabilization by the protein of one specific conformation from a pool of conformations reflecting the inherent flexibility of the RNA (Williamson 2000;Leulliot and Varani 2001). One example of protein-assisted RNA folding according to the induced fit mechanism is the binding of the 3Ј UTR of U1A pre-mRNA to the U1A protein (Oubridge et al 1994;Avis et al 1996;Gubser and Varani 1996;Varani et al 2000). Previous studies proposed a protein-assisted RNA folding for the K-turn motif (Matsumura et al 2003;Goody et al 2004).…”

Section: Introductionmentioning

confidence: 99%

The snRNP 15.5K protein folds its cognate K-turn RNA: A combined theoretical and biochemical study

Cojocaru¹,

Nottrott²,

Klement³

et al. 2005

RNA

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The human 15.5K protein binds to the 5 stem-loop of U4 snRNA, promotes the assembly of the spliceosomal U4/U6 snRNP, and is required for the recruitment of the 61K protein and the 20/60/90K protein complex to the U4 snRNA. In the crystallographic structure of the 15.5K-U4 snRNA complex, the conformation of the RNA corresponds to the family of kink-turn (K-turn) structural motifs. We simulated the complex and the free RNA, showing how the protein binding and the intrinsic flexibility contribute to the RNA folding process. We found that the RNA is significantly more flexible in the absence of the 15.5K protein. Conformational transitions such as the interconversion between alternative purine stacking schemes, the loss of G-A base pairs, and the opening of the K-turn occur only in the free RNA. Furthermore, the stability of one canonical G-C base pair is influenced both by the binding of the 15.5K protein and the nature of the adjacent structural element in the RNA. We performed chemical RNA modification experiments and observed that the free RNA lacks secondary structure elements, a result in excellent agreement with the simulations. Based on these observations, we propose a protein-assisted RNA folding mechanism in which the RNA intrinsic flexibility functions as a catalyst.

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Structure of the Polyadenylation Regulatory Element of the Human U1A Pre-mRNA 3‘-Untranslated Region and Interaction with the U1A Protein

Cited by 75 publications

References 48 publications

Recognition of the Core RNA Promoter for Minus-Strand RNA Synthesis by the Replicases of Brome Mosaic Virus and Cucumber Mosaic Virus

Recognition of the Core RNA Promoter for Minus-Strand RNA Synthesis by the Replicases of Brome Mosaic Virus and Cucumber Mosaic Virus

NMR studies of dynamics in RNA and DNA by ¹³C relaxation

The snRNP 15.5K protein folds its cognate K-turn RNA: A combined theoretical and biochemical study

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Structure of the Polyadenylation Regulatory Element of the Human U1A Pre-mRNA 3‘-Untranslated Region and Interaction with the U1A Protein

Cited by 75 publications

References 48 publications

Recognition of the Core RNA Promoter for Minus-Strand RNA Synthesis by the Replicases of Brome Mosaic Virus and Cucumber Mosaic Virus

Recognition of the Core RNA Promoter for Minus-Strand RNA Synthesis by the Replicases of Brome Mosaic Virus and Cucumber Mosaic Virus

NMR studies of dynamics in RNA and DNA by 13C relaxation

The snRNP 15.5K protein folds its cognate K-turn RNA: A combined theoretical and biochemical study

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NMR studies of dynamics in RNA and DNA by ¹³C relaxation